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The inflammatory cytokines interleukin-1␤ and tumor necrosis factor-␣ (TNF-␣) have been identified as mediators of several forms of neurodegeneration in the brain. However, they can produce either deleterious or beneficial effects on neuronal function. We investigated the effects of these cytokines on neuronal death caused by exposure of mouse organotypic hippocampal slice cultures to toxic concentrations of AMPA. Either potentiation of excitotoxicity or neuroprotection was observed, depending on the concentration of the cytokines and the timing of exposure. A relatively high concentration of mouse recombinant TNF-␣ (10 ng/ml) enhanced excitotoxicity when the cultures were simultaneously exposed to AMPA and to this cytokine. Decreasing the concentration of TNF-␣ to 1 ng/ml resulted in neuroprotection against AMPA-induced neuronal death independently on the application protocol. By using TNF-␣ receptor (TNFR) knock-out mice, we demonstrated that the potentiation of AMPA-induced toxicity by TNF-␣ involves TNF receptor-1, whereas the neuroprotective effect is mediated by TNF receptor-2. AMPA exposure was associated with activation and proliferation of microglia as assessed by macrophage antigen-1 and bromodeoxyuridine immunohistochemistry, suggesting a functional recruitment of cytokineproducing cells at sites of neurodegeneration. Together, these findings are relevant for understanding the role of proinflammatory cytokines and microglia activation in acute and chronic excitotoxic conditions.

Tumor necrosis factor (TNF)-␣ has been reported to modulate brain injury, but remarkably, little is known about its effects on neurogenesis. We report that TNF-␣ strongly influences survival, proliferation, and neuronal differentiation in cultured subventricular zone (SVZ) neural stem/progenitor cells derived from the neonatal P1-3 C57BL/6 mice. By using single-cell calcium imaging, we

The manipulation of endogenous stem cell populations from the subventricular zone (SVZ), a neurogenic niche, creates an opportunity to induce neurogenesis and influence brain regenerative capacities in the adult brain. Herein, we demonstrate the ability of polyelectrolyte nanoparticles to induce neurogenesis exclusively after being internalized by SVZ stem cells. The nanoparticles are not cytotoxic for concentrations equal or below 10 μg/mL. The internalization process is rapid, and nanoparticles escape endosomal fate in a few hours. Retinoic acid-loaded nanoparticles increase the number of neuronal nuclear protein (NeuN)-positive neurons and functional neurons responding to depolarization with KCl and expressing NMDA receptor subunit type 1 (NR1). These nanoparticles offer an opportunity for in vivo delivery of proneurogenic factors and neurodegenerative disease treatment.

Modulation of the subventricular zone (SVZ) neurogenic niche can enhance brain repair in several disorders including Parkinson's disease (PD). Herein, we used biocompatible and traceable polymeric nanoparticles (NPs) containing perfluoro-1,5-crown ether (PFCE) and coated with protamine sulfate to complex microRNA-124 (miR-124), a neuronal fate determinant. The ability of NPs to efficiently deliver miR-124 and prompt SVZ neurogenesis and brain repair in PD was evaluated. In vitro, miR-124 NPs were efficiently internalized by neural stem/progenitors cells and neuroblasts and promoted their neuronal commitment and maturation. The expression of Sox9 and Jagged1, two miR-124 targets and stemness-related genes, were also decreased upon miR-124 NP treatment. In vivo, the intracerebral administration of miR-124 NPs increased the number of migrating neuroblasts that reached the granule cell layer of the olfactory bulb, both in healthy and in a 6-hydroxydopamine (6-OHDA) mouse model for PD.MiR-124 NPs were also able to induce migration of neurons into the lesioned striatum of 6-OHDA-treated mice. Most importantly, miR-124 NPs proved to ameliorate motor symptoms of 6-OHDA mice, monitored by the apomorphine-induced rotation test.Altogether, we provide clear evidences to support the use of miR-124 NPs as a new therapeutic approach to boost endogenous brain repair mechanisms in a setting of neurodegeneration.

Summary:Purpose: Cytokines and related inflammatory mediators are rapidly synthesized in the brain during seizures. We previously found that intracerebral administration of interleukin-1 (IL-1)-β has proconvulsant effects, whereas its endogenous receptor antagonist (IL-1Ra) mediates potent anticonvulsant actions in various models of limbic seizures. In this study, we investigated whether seizures can be effectively inhibited by blocking the brain production of IL-1β, by using selective inhibitors of interleukin-converting enzyme (ICE/caspase-1) or through caspase-1 gene deletion.Methods: Caspase-1 was selectively blocked by using pralnacasan or VX-765. IL-1β release was induced in mouse organotypic hippocampal slice cultures by proinflammatory stimuli [lipopolysaccaride (LPS) + adenosine triphosphate (ATP)] and measured with enzyme-linked immunosorbent assay (ELISA). IL-1β production during seizures was measured in the rat hippocampus by Western blot. Seizures were induced in freely moving mice and rats by intrahippocampal injection of kainic acid and recorded by EEG analysis.Results: Caspase-1 inhibition reduced the release of IL-1β in organotypic slices exposed to LPS+ATP. Administration of pralnacasan (intracerebroventricular, 50 μg) or VX-765 (intraperitoneal, 25-200 mg/kg) to rats blocked seizure-induced production of IL-1β in the hippocampus, and resulted in a twofold delay in seizure onset and 50% reduction in seizure duration. Mice with caspase-1 gene deletion showed a 70% reduction in seizures and an approximate fourfold delay in their onset.Conclusions: Inhibition of caspase-1 represents an effective and novel anticonvulsive strategy, which acts by selectively reducing the brain availability of IL-1β.

BackgroundHistamine is commonly acknowledged as an inflammatory mediator in peripheral tissues, leaving its role in brain immune responses scarcely studied. Therefore, our aim was to uncover the cellular and molecular mechanisms elicited by this molecule and its receptors in microglia-induced inflammation by evaluating cell migration and inflammatory mediator release.MethodsFirstly, we detected the expression of all known histamine receptor subtypes (H1R, H2R, H3R and H4R), using a murine microglial cell line and primary microglia cell cultures from rat cortex, by real-time PCR analysis, immunocytochemistry and Western blotting. Then, we evaluated the role of histamine in microglial cell motility by performing scratch wound assays. Results were further confirmed using murine cortex explants. Finally, interleukin-1beta (IL-1β) and tumor necrosis factor-alpha (TNF-α) levels were evaluated by ELISA measurements to determine the role of histamine on the release of these inflammatory mediators.ResultsAfter 12 h of treatment, 100 μM histamine and 10 μg/ml histamine-loaded poly (lactic-co-glycolic acid) microparticles significantly stimulated microglia motility via H4R activation. In addition, migration involves α5β1 integrins, and p38 and Akt signaling pathways. Migration of microglial cells was also enhanced in the presence of lipopolysaccharide (LPS, 100 ng/ml), used as a positive control. Importantly, histamine inhibited LPS-stimulated migration via H4R activation. Histamine or H4R agonist also inhibited LPS-induced IL-1β release in both N9 microglia cell line and hippocampal organotypic slice cultures.ConclusionsTo our knowledge, we are the first to show a dual role of histamine in the modulation of microglial inflammatory responses. Altogether, our data suggest that histamine per se triggers microglia motility, whereas histamine impedes LPS-induced microglia migration and IL-1β release. This last datum assigns a new putative anti-inflammatory role for histamine, acting via H4R to restrain exacerbated microglial responses under inflammatory challenge, which could have strong repercussions in the treatment of CNS disorders accompanied by microglia-derived inflammation.

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